This invention relates to liquid scintillation counting. In particular the invention is concerned with a system for monitoring and measuring the color of samples in a liquid scintillation counter. The system is directed to determining whether the color in a liquid scintillation sample is sufficiently intense such that direct activity, namely disintegrations per minute (DPM) of the sample can be determined from a conventional chemical quench curve.
It is known that fewer photons leave the vial of a quenched liquid scintillation sample relative to photons leaving an unquenched sample. Three primary quenching processes are known: chemical, color, and absorption. Each of these processes interferes with one of the energy exchange processes required for the production of light and its detection by a photomultiplier tube.
The effect of quenching can be described in relation to the following energy transfer processes which take place in a liquid scintillation cocktail.
______________________________________ 1. .sup.--B + .sup.--S .fwdarw. S + B electronic excitation of the solvent 2. .sup.--S + .sup.--F .fwdarw. S + F electronic excitation of the Fluor 3. .sup.--F .fwdarw. F + hV photon emission by the Fluor 4. hV + PMT .fwdarw. Ep photoelectric electron emission by the PMT ______________________________________
A beta particle, B, with kinetic energy interacts with a solvent molecule, S, in the cocktail causing its electronic excitation, S; step 1. The excess electronic energy of the solvent can be passed to a fluor molecule, F, causing its electronic excitation, F; step 2. The excited fluor molecule can emit a photon, hV; step 3, and return to the electronic ground state. This photon, after leaving the vial, can interact with the cathode of a photomultiplier tube and produce a photoelectric electron. This signal is amplified and represents the "observation of a radionuclide decay event."
Quench refers to any process which interferes with the energy exchange reactions represented in the above steps. Absorption quench refers to processes which interfere with step 1. In other words, a beta particle is prevented by absorption from reacting with a solvent molecule. Chemical quench refers to interference either with excitation of F, step 2, or via internal energy mode decay which prevents production of hV in step 3. Some molecule other than the solvent is excited by the beta particle, thereby preventing solvent excitation. Color quench refers to any chemical which absorbs hV produced by the fluor so that light does not reach the PMT in step 4.
The wavelength of the emitted light depends upon the specific fluor. This generally lies between 380-430 nm; however, the emission bands are broad and may extend beyond these wavelengths. Any substance absorbing energy in the stated wavelength range decreases the number of photons leaving the sample vial. Where the DPM in the sample is required a quench curve is used. This requires the counting efficiency, E, of the sample as defined by: ##EQU1## where the counts per minute (CPM) is observed for a sample having DPM disintegrations per minute. To obtain DPM for an unknown, CPM is measured directly by a liquid scintillation counter while E is obtained from a quench curve previously prepared. A quench curve relates nuclide counting efficiency, E, to some quench monitor, for instance the H# in Horrocks (U.S. Pat. No. 4,075,480). A quench curve is developed from a set of quenched standards, each containing the same number of DPM's of the same radionuclide, but containing a different quantity of a chemical quench agent.
With the availability of the quench curve, the DPM of an unknown sample is obtained by measurement of the H# and CPM of the sample. The counting efficiency is obtained from the H# and quench curve.
This gives accurate values for DPM provided that only chemical quench is present. Should color quench be present, then a chemical quench curve does not always recover DPM correctly, since there is a difference between chemical and color quench curves in terms of H#. As the level of quench increases, the difference between the chemical and color quench curves increases, thus leading to increased percentage error in DPM.
Different quench curves are thus required for chemically quenched standards and for a set of color quenched standards. For measuring an unknown sample it would be necessary to know beforehand which quench curve to use. Furthermore, if an unknown contained both chemical and color quenching agents or a color agent which absorbed at a wavelength different from the agent used for the quench curve standard, then the available curves would not provide the correct answer. Accordingly, a variety of quench curves is needed to answer correctly for a variety of chromophores at a variety of quench levels.
Some limited solutions for color quench correction have been suggested. The suggested color quench curve corrections have been for specific systems with one exception. Specific systems are those with no variations between the standard and the unknown regarding the cocktail, chromophore or level of quench present.
Also, quench monitors with limitations have been reported; none however, has been placed on an automatic liquid scintillation counter.
One prior art approach depends upon correlations between counting efficiency and spectroscopic parameters such as absorbance or wavelength of the absorber. A second approach depends upon the development of and analysis of four different quench curves which are functions of two different quench monitors for the two types of quench, color and chemical. The third approach involves some variation on the concept of lesser pulse height analysis. A fourth approach includes isolated internal standards and spectral analysis Additionally other investigators have sought to solve the problem in effect by either establishing color quench curves and measuring unknowns from them or by decolorizing the system through the use of chemical oxidizing agents.
There has been limited success with the spectroscopy approach in trying to correlate the primary wavelength of absorption of the chromophore with the counting efficiency of the sample. A multicomponent analysis approach to the use of the Beer-Lambert law correlating absorbance and the counting efficiency of the sample has also been attempted. Difficulties with these methods include the numerous measurements and the awkwardness of automation.
Using the four quench curve approach, two quench curves are developed for pure chemical quench and two for pure color quench. For an unknown having pure chemical quench, the two quench curves for chemical quench would give the same values for the counting efficiency while the two color curves would not. Should the unknown be pure color quenched, then the two color quench curves would give the same efficiency while the chemical quench curves would not. This scheme is not useful if both chemical and color quench are present. An advantage of this method however is that it provides a built-in quench monitor.
With the lesser pulse height analysis technique, the pulses from two photomultiplier tubes are examined separately. This allows mathematical analysis to combine advantageously the results of summation counting from both tubes with single tube results either in or out of coincidence.
In general, if light is generated at a point, in a sample, then one path, to one photomultiplier tube is shorter than a second path, to a second photomultiplier tube. This difference is important, since, if color is present, absorbance, As, is path length dependent, L. Absorbance is also concentration, C, and chromophore dependent, A , according to the Beer-Lambert law: EQU As=ALC
Assuming that the same number of photons leave along both paths, then fewer photons will arrive at one tube than the other tube in accordance with the Beer-Lambert law because the one path is longer than the other path. The pulse arriving at the one tube will be of lesser intensity. In general, for each decay event in the presence of color, the intensities of the two pulses arriving at the respective tubes will be different. Laney (U.S. Pat. No. 3,725,657) and Jordan (U.S. Pat. No. 4,292,520) disclose using the lesser intense pulse to monitor counting efficiency.
Here, the separated color and chemical quench curves which occur if summed coincidence is used, become one curve if the lesser pulse is used. In other words, the color effect is not monitored by the lesser pulse, since it is lost. This is both advantageous and disadvantageous. One advantageous aspect is that only one quench curve per radionuclide is required to obtain counting efficiencies for both color and chemically quenched samples. The technique however does not solve the color correction problem in a general sense, since the chemical and color quench curves split apart again at higher quench levels even if lesser pulse height analysis is used. The lesser pulse height technique is also not capable of providing a color monitor since the technique is insensitive to color.
With an isolated internal standard technique a "sealed internal standard" in a pyrex tube with a small internal diameter is prepared. This standard contains a sample of the radionuclide to be measured in an unknown. Preparation of sealed internal standards for each radionuclide of interest is required. The mechanical placement of the internal standard into a sample vial, its removal and cleaning prior to use in the next sample is not conducive to an automated operation.
No combination of color monitor and color correction has been developed together and incorporated onto an automated liquid scintillation counter in any of the methods discussed above.
It is an object of the present invention to provide a monitor to determine whether a given chemical quench curve can be applied accurately to a given sample.